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Article Dans Une Revue Biochemical Journal Année : 2008

Characterisation of cytochrome bo3 activity in a native-like surface-tethered membrane

Sophie A. Weiss
  • Fonction : Auteur
Richard J Bushby
  • Fonction : Auteur
Stephen D. Evans
  • Fonction : Auteur
Pjf Henderson
  • Fonction : Auteur

Résumé

We have developed a simple native-like surface-tethered membrane system to investigate the activity of cytochrome bo3 (cbo3), a terminal oxidase in Escherichia coli. The tethered membranes consist of E. coli inner membrane extracts mixed with additional E. coli lipids containing various amounts of the cbo3 substrate ubiquinol-10 (UQ-10). Tethered membranes are formed by self assembly from vesicles onto gold electrodes functionalised with cholesterol derivatives. Cytochrome bo3 activity was monitored using cyclic voltammetry with electron transfer to cbo3 mediated by UQ-10. The apparent KM for oxygen with this system is 1.1±0.4 μM, in good agreement with literature values for whole cell experiments and for purified cbo3. Increasing the concentration of lipophilic UQ-10 in the membrane leads to an increase in cbo3 activity. The activity of cbo3 with long chain ubiquinones appears to be different to previous reports using short chain substrate analogues such as UQ-1 in that typical Michaelis Menten kinetics are not observed using UQ-10. This native-like membrane model thus provides new insights into the interaction of transmembrane enzymes with hydrophobic substrates which contrasts with studies using hydrophilic UQ analogues.

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Dates et versions

hal-00479061 , version 1 (30-04-2010)

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Sophie A. Weiss, Richard J Bushby, Stephen D. Evans, Pjf Henderson, Lars J C Jeuken. Characterisation of cytochrome bo3 activity in a native-like surface-tethered membrane. Biochemical Journal, 2008, 417 (2), pp.555-560. ⟨10.1042/BJ20081345⟩. ⟨hal-00479061⟩

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